Investigation of DNA polymorphisms in SMAD genes for genetic predisposition to diabetic nephropathy in patients with type 1 diabetes mellitus

Abstract

Aims/hypothesis SMAD proteins are involved in multiple signalling pathways and are key modulators of gene expression. We hypothesised that genetic variation in selected SMAD genes contributes to susceptibility to diabetic nephropathy. Methods We selected 13 haplotype tag (ht) single nucleotide polymorphisms (SNPs) from 67 variants identified by resequencing the SMAD2 and SMAD3 genes. For SMAD1, SMAD4 and SMAD5 genes, genotype data were downloaded for 217 SNPs from Phase II of the International HapMap project. Of these, 85 SNPs met our inclusion criteria, resulting in the selection of 13 tag SNPs for further investigation. A case-control approach was employed, using 267 nephropathic patients and 442 controls with type 1 diabetes from Ireland. Two further populations (totalling 1,407 patients, 2,238 controls) were genotyped to validate initial findings. Genotyping was conducted using iPLEX, TaqMan and gel electrophoresis. Results The distribution of genotypes was in HardyWeinberg equilibrium. Analysis by the χ 2 test of genotype and allele frequencies in patients versus controls in the Irish population (n=709) revealed evidence for the association of one allele at 5% level of significance (rs10515478, p uncorrected =0.006; p corrected =0.04). This finding represents a relatively small difference in allele frequency of 6.4% in the patient group compared with 10.7% in the control group; this difference was not supported in subsequent investigations using DNA from European individuals with similar phenotypic characteristics. Conclusions/interpretation We selected an appropriate subset of variants for the investigation of common genetic risk factors and assessed SMAD1 to SMAD5 genes for association with diabetic nephropathy. We conclude that common polymorphisms in these genes do not strongly influence genetic susceptibility to diabetic nephropathy in white individuals with type 1 diabetes mellitus.

Resequencing of the CCL5 and CCR5 genes and investigation of variants for association with diabetic nephropathy

“…Our case-control collection was selected using strict phenotypic criteria and has been previously used to investigate association with DN. 18,20,21 Greater than 95% completion was achieved for all genotyped SNPs. Contingency table w 2 tests were used to compare genotype and allele frequencies between cases and controls.…”

“…18 Briefly, cases have type 1 diabetes with DN, while controls are type 1 diabetic patients without nephropathy. The average age at recruitment of cases and controls was 48.7 (s.d.…”

“…Bidirectional sequencing, screening for variants, haplotype tag singlenucleotide polymorphism (htSNP) selection and genotyping were performed as previously described. 18 The resequenced data have been submitted to GenBank as (a) GQ504011 C-C chemokine ligand 5 (CCL5) gene, promoter region and complete cds, and as (b) GQ917109 C-C chemokine receptor type 5 (CCR5) gene and complete cds. Novel SNPs have been submitted to dbSNP (Table 1).…”

A GREM1 Gene Variant Associates with Diabetic Nephropathy

“…Individuals have been described previously in case-control association studies investigating diabetic nephropathy 2 ; characteristics are presented in Table 1. Resequencing data were submitted to GenBank as (1) DQ100069 Gremlin 1 homolog, cysteine knot superfamily (X. laevis) (GREM1) 5Ј upstream, promoter and exon 1, genomic sequence; and (2) DQ100070 Gremlin 1 homolog, cysteine knot superfamily (X. laevis) (GREM1) coding region and 3Ј downstream genomic sequence.…”

“…1 The development of larger carefully phenotyped case-control collections now permits appropriately designed studies to be conducted with improved statistical power. 2,3 Gremlin is implicated in several developmental pathways 4,5 and has become increasingly recognized as an important contributor to renal disease. 6 -8 Gremlin influences cell growth and differentiation, 9 particularly through bone morphogenic protein and transforming growth factor-␤-mediated processes.…”